Morphological and life-history trait plasticity of two Daphnia species induced by fish kairomones.

Daphnia can avoid predation by sensing fish kairomones and producing inducible defenses by altering the phenotype. In this study, the results showed that the morphological and life-history strategies of two Daphnia species (Daphnia pulex and Daphnia sinensis) exposed to Aristichthys nobilis kairomones. In the presence of fish kairomones, the two Daphnia species exhibited significantly smaller body length at maturity, smaller body length of offspring at the 10th instar, and longer relative tail spine of offspring. Nevertheless, other morphological and life-history traits of the two Daphnia species differed. D. pulex showed a significantly longer relative tail spine length and earlier age at maturity after exposure to fish kairomones. The total offspring number of D. sinensis exposed to fish kairomones was significantly higher than that of the control group, whereas that of D. pulex was significantly lower. These results suggest that the two Daphnia species have different inducible defense strategies (e.g., morphological and life-history traits) during prolonged exposure to A. nobilis kairomones, and their offspring also develop morphological defenses to avoid predation. It will provide reference for further exploring the adaptive evolution of Daphnia morphology and life-history traits in the presence of planktivorous fish.

TREM2 receptor protects against complement-mediated synaptic loss by binding to complement C1q during neurodegeneration.

Triggering receptor expressed on myeloid cells 2 (TREM2) is strongly linked to Alzheimer's disease (AD) risk, but its functions are not fully understood. Here, we found that TREM2 specifically attenuated the activation of classical complement cascade via high-affinity binding to its initiator C1q. In the human AD brains, the formation of TREM2-C1q complexes was detected, and the increased density of the complexes was associated with lower deposition of C3 but higher amounts of synaptic proteins. In mice expressing mutant human tau, Trem2 haploinsufficiency increased complement-mediated microglial engulfment of synapses and accelerated synaptic loss. Administration of a 41-amino-acid TREM2 peptide, which we identified to be responsible for TREM2 binding to C1q, rescued synaptic impairments in AD mouse models. We thus demonstrate a critical role for microglial TREM2 in restricting complement-mediated synaptic elimination during neurodegeneration, providing mechanistic insights into the protective roles of TREM2 against AD pathogenesis.

ß2-Microglobulin coaggregates with Aß and contributes to amyloid pathology and cognitive deficits in Alzheimer's disease model mice.

Extensive studies indicate that ß-amyloid (Aß) aggregation is pivotal for Alzheimer's disease (AD) progression; however, cumulative evidence suggests that Aß itself is not sufficient to trigger AD-associated degeneration, and whether other additional pathological factors drive AD pathogenesis remains unclear. Here, we characterize pathogenic aggregates composed of ß2-microglobulin (ß2M) and Aß that trigger neurodegeneration in AD. ß2M, a component of major histocompatibility complex class I (MHC class I), is upregulated in the brains of individuals with AD and constitutes the amyloid plaque core. Elevation of ß2M aggravates amyloid pathology independent of MHC class I, and coaggregation with ß2M is essential for Aß neurotoxicity. B2m genetic ablation abrogates amyloid spreading and cognitive deficits in AD mice. Antisense oligonucleotide- or monoclonal antibody-mediated ß2M depletion mitigates AD-associated neuropathology, and inhibition of ß2M-Aß coaggregation with a ß2M-based blocking peptide ameliorates amyloid pathology and cognitive deficits in AD mice. Our findings identify ß2M as an essential factor for Aß neurotoxicity and a potential target for treating AD.

ß2-microglobulin functions as an endogenous NMDAR antagonist to impair synaptic function.

Down syndrome (DS) is a neurological disorder with multiple immune-related symptoms; however, crosstalk between the CNS and peripheral immune system remains unexplored. Using parabiosis and plasma infusion, we found that blood-borne factors drive synaptic deficits in DS. Proteomic analysis revealed elevation of ß2-microglobulin (B2M), a major histocompatibility complex class I (MHC-I) component, in human DS plasma. Systemic administration of B2M in wild-type mice led to synaptic and memory defects similar to those observed in DS mice. Moreover, genetic ablation of B2m or systemic administration of an anti-B2M antibody counteracts synaptic impairments in DS mice. Mechanistically, we demonstrate that B2M antagonizes NMDA receptor (NMDAR) function through interactions with the GluN1-S2 loop; blocking B2M-NMDAR interactions using competitive peptides restores NMDAR-dependent synaptic function. Our findings identify B2M as an endogenous NMDAR antagonist and reveal a pathophysiological role for circulating B2M in NMDAR dysfunction in DS and related cognitive disorders.

SNX14 deficiency-induced defective axonal mitochondrial transport in Purkinje cells underlies cerebellar ataxia and can be reversed by valproate.

Loss-of-function mutations in sorting nexin 14 (SNX14) cause autosomal recessive spinocerebellar ataxia 20, which is a form of early-onset cerebellar ataxia that lacks molecular mechanisms and mouse models. We generated Snx14-deficient mouse models and observed severe motor deficits and cell-autonomous Purkinje cell degeneration. SNX14 deficiency disrupted microtubule organization and mitochondrial transport in axons by destabilizing the microtubule-severing enzyme spastin, which is implicated in dominant hereditary spastic paraplegia with cerebellar ataxia, and compromised axonal integrity and mitochondrial function. Axonal transport disruption and mitochondrial dysfunction further led to degeneration of high-energy-demanding Purkinje cells, which resulted in the pathogenesis of cerebellar ataxia. The antiepileptic drug valproate ameliorated motor deficits and cerebellar degeneration in Snx14-deficient mice via the restoration of mitochondrial transport and function in Purkinje cells. Our study revealed an unprecedented role for SNX14-dependent axonal transport in cerebellar ataxia, demonstrated the convergence of SNX14 and spastin in mitochondrial dysfunction, and suggested valproate as a potential therapeutic agent.

Imaging of individual transcripts by amplification-based single-molecule fluorescence in situ hybridization.

An amplification-based single-molecule fluorescence in situ hybridization (asmFISH) assay is introduced that exploits improved probe design for highly specific imaging of individual transcripts in fixed cells and tissues. In this method, a pair of DNA ligation probes are ligated on RNA templates upon specific hybridization, followed by probe circularization based on enzymatic DNA ligation and rolling circle amplification for signal boosting. The method is more efficient and specific than the padlock probe assay for detection of the same RNA molecules and discrimination of single nucleotide polymorphisms. Moreover, asmFISH is a versatile method which can be applied not only to cultured cells, but also to fresh frozen and formalin-fixed, paraffin-embedded tissue sections.

Visualization of individual microRNA molecules in fixed cells and tissues using target-primed padlock probe assay.

MicroRNAs (miRNAs) are key regulators of gene expression at the posttranscriptional level. Precisely profiling of miRNA expression will help us to better understand their roles in normal and diseased cells and tissues. Here we describe in situ miRNA detection by padlock probing and miRNA target-primed rolling circle amplification. We optimized our protocol and showed it can be applied to both fixed cells and tissue sections. The method can be used in basic research and potentially in clinical diagnostics in the future.

Novel KCNJ10 Compound Heterozygous Mutations Causing EAST/SeSAME-Like Syndrome Compromise Potassium Channel Function.

Inwardly rectifying K+ channel 4.1 (Kir4.1), encoded by KCNJ10, is a member of the inwardly rectifying potassium channel family. In the brain, Kir4.1 is predominant in astrocytic glia and accounts for the spatial buffering of K+ released by neurons during action potential propagation. A number of studies have shown that mutations in KCNJ10 are associated with SeSAME/EAST syndrome, which is characterized by seizures, ataxia, sensorineural deafness, and electrolyte imbalance. Herein, we identified two siblings presenting with seizures and motor delays in one outbred kindred. Customized targeted-exome sequencing showed that both affected siblings are compound heterozygous for two KCNJ10 missense mutations (NM_002241.4: c.601G > A: p.A201T and c.626T > C: p.I209T). Prediction tools suggested that both amino acid substitutions were deleterious or disease causing. Further functional studies showed that Chinese hamster ovary (CHO) cells expressing either A201T and/or I209T Kir4.1 channels exhibited lower K+ currents, indicating compromised Kir4.1 biological function. Intriguingly, the A201T but not I209T mutation decreased total and cell surface Kir4.1 levels. Kir4.1 channels with the A201T mutation were unstable and degraded through lysosomal pathway. In conclusion, these data indicated that both A201T and I209T mutations disrupt Kir4.1 activity and are the cause of SeSAME/EAST-like syndrome in the siblings.

The Retromer Complex and Sorting Nexins in Neurodegenerative Diseases.

The retromer complex and associated sorting nexins (SNXs) comprise a critical trafficking machinery which mediates endosomal protein sorting. Retromer and/or SNX dysfunction has been linked to several neurodegenerative diseases including Alzheimer's disease (AD), Parkinson's disease (PD), and Down's syndrome (DS). In AD, deficiency of the retromer complex or its cargo proteins impairs endosomal trafficking of amyloid precursor protein (APP), resulting in the overproduction of ß-amyloid (Aß). Several SNX components directly interact with APP or APP-cleaving enzymes (ß- and γ-secretases) to regulate amyloidogenic APP processing and Aß generation. In addition, PD-linked mutations in retromer components cause mistrafficking of retromer cargo proteins and mitochondrial dysfunction, and dysregulation retromer-mediated trafficking has been considered as an important cause of hereditary spastic paraplegia (HSP) and neuronal ceroid lipofuscinoses (NCLs). Moreover, SNX27 deficiency is an important contributor for synaptic and cognitive impairment in DS. Here we review recent findings describing the retromer complex and/or SNXs-mediated endosomal sorting in neurodegenerative disorders.

One-Step Method to Prepare PLLA Porous Microspheres in a High-Voltage Electrostatic Anti-Solvent Process.

A one-step method using a high-voltage electrostatic anti-solvent process was employed to fabricate poly-l-lactide (PLLA) porous microspheres (PMs). To address the simplification and control of the preparation process, a 24 full factorial experiment was performed to optimize the operating process and analyze the effect of the factors on the morphology and aerodynamic properties of the PLLA PMs, and various characterization tests were also performed. The resulting PLLA PMs exhibited an even and porous morphology with a density less than 0.4 g/cm³, a geometric mean diameter (Dg) of 10-30 µm, an aerodynamic diameter (Da) of 1-5 µm, a fine particle fraction (FPF) of 56.3%, and a porosity of 76.2%, meeting the requirements for pulmonary drug delivery. The physicochemical characterizations reveal that no significant chemical change occurred in the PLLA during the process. An investigation of its in vitro cytotoxicity and pulmonary toxicity shows no obvious toxic response, indicating good biosafety. This study indicates that the one-step method using a high-voltage electrostatic anti-solvent process has great potential in developing an inhalable drug carrier for pulmonary drug delivery.

A Rab8 guanine nucleotide exchange factor-effector interaction network regulates primary ciliogenesis.

Primary cilia are microtubule-based solitary membrane projections on the cell surface that play important roles in signaling and development. Recent studies have demonstrated that polarized vesicular trafficking involving the small GTPase Rab8 and its guanine nucleotide exchange factor Rabin8 is essential for primary ciliogenesis. In this study, we show that a highly conserved region of Rabin8 is pivotal for its activation as a guanine nucleotide exchange factor for Rab8. In addition, in its activated conformation, Rabin8 interacts with Sec15, a subunit of the exocyst and downstream effector of Rab8. Expression of constitutively activated Rab8 promotes the association of Sec15 with Rabin8. Using immunofluorescence microscopy, we found that Sec15 co-localized with Rab8 along the primary cilium. Inhibition of Sec15 function in cells led to defects in primary ciliogenesis. The Rabin8-Rab8-Sec15 interaction may couple the activation of Rab8 to the recruitment of the Rab8 effector and is involved in the regulation of vesicular trafficking for primary cilium formation.

[Evaluation of a model of temporomandibular disorders established by transzygomatic arch traction of the mandibular ramus in rabbits].

OBJECTIVE: To evaluate a model of temporomandibular disorders established by transzygomatic arch traction of the mandibular ramus in rabbits. METHODS: Fifteen adult New Zealand rabbits were subjected to traction in the postero-superior direction unilaterally using elastic force and six rabbits used as the control. Histopathologic change of the disc, joint space and cartilage was observed through Hematoxylin and Eosin staining. RESULTS: Anterior disc displacement or disc deformity in four experimental rabbits was observed on the traction side 2 weeks after operation. At 4 weeks, fibrous adhesions in joint compartment were found in five experimental rabbits. The condyles or articular eminences of some experimental rabbits showed irregularities on the cartilage surface. In the 6 th week, bad disc deformity in four rabbits and severe fibrous adhesions in five rabbits was observed on the traction side, and subchondralbone and calcified cartilage became irregular. In control group, All articular structures were normal. CONCLUSIONS: A animal model of temporomandibular disorders can be established by transzygomatic arch traction of the mandible.

[The distribution of collagen I, II, X and alkaline phosphatase in the development of condylar cartilage of fetal mouse mandible].

OBJECTIVE: The purpose of this study was to investigate the distribution of collagen I, II , X, alkaline phosphatase (ALP) and their roles during initiation of condylar cartilage of the fetal mouse. METHODS: Coronary sections of mandible of mouse embryo aged from 14th to 18th day were studied under light microscope after stained by immunohistochemical method with antibody of types I, II, X collagen and ALP. RESULTS: On the 14th day of mouse embryo, it was found that mesenchymal cells condensation continuous with the periosteum. Type I collagen and ALP were positive behind the terminal of the ossifying mandibular periosteum where future condylar will form. On the 15th day, positive staining for types I, II collagen was found in mesenchymal cells around hypertrophic cells and type X collagen was detected in hypertrophic cells. ALP was positive in both mesenchymal cells and hypertrophic cells. On the 16th day, type I collagen was observed from periosteal osteogenic cells and mesenchymal cells of the fibrous cell layer to the upper hypertrophic cell layer while Type II collagen was restricted from the lower polymorphic cell layer to the bottom of the hypertropic cell layer. Type X collagen was positive in the hypertrophic cell layer. ALP was positive in periosteal osteogenic cells and hypertrophic chondral cells, but not in the polymorphic cell layer. CONCLUSION: Development of condylar cartilage is different from that of limb bone. Types I, II, X collagen are expressed in the condylar chondrocyte on the early stage of endochondral ossification. The histology evidence supports the conjecture that condylar cartilage is derived from differentiated mesenchymal cells of the preperiosteum or periosteum of the mandible where ALP is positively expressed.